#knitr::opts_chunk$set(message = FALSE, echo = FALSE)
library(Matrix)
library(genomation)
## Loading required package: grid
library(matrixStats)
library(knitr)
library(ggplot2)
library(ArchR)
## Loading required package: SummarizedExperiment
## Loading required package: GenomicRanges
## Loading required package: stats4
## Loading required package: BiocGenerics
## Loading required package: parallel
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## Attaching package: 'BiocGenerics'
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##     'browseVignettes()'. To cite Bioconductor, see
##     'citation("Biobase")', and for packages 'citation("pkgname")'.
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## Attaching package: 'Biobase'
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## Loading required package: rhdf5
## Loading required package: magrittr

Data was preprocessed by Tiffany using the Cell Ranger Pipeline: https://support.10xgenomics.com/single-cell-atac/software/pipelines/latest/algorithms/overview

Based on marker genes, plots below,

  • Cluster 1-3 express HBC markers
  • Cluster 4 seems to be neuronal (Omp, Cnga2 only expressed in cl4). Seeems split in 2, possibly inhibitory and excitatory?
  • Cluster 5 expresses SUS markers (Cyp2g1).
  • Cluster 6 doesn’t really seem to be consistent and doesnt really express anything so may be low quality cells.
## Setting default genome to Mm10.
## Using GeneAnnotation set by addArchRGenome(Mm10)!
## Using GeneAnnotation set by addArchRGenome(Mm10)!
## Cleaning Temporary Files
## 2021-01-28 22:54:29 : Batch Execution w/ safelapply!, 0 mins elapsed.
## (s1Inj : 1 of 6) Checking if completed file exists!
## 2021-01-28 22:54:29 : (s1Inj : 1 of 6) Arrow Exists! Marking as completed since force = FALSE!, 0.001 mins elapsed.
## (s3Inj : 2 of 6) Checking if completed file exists!
## 2021-01-28 22:54:29 : (s3Inj : 2 of 6) Arrow Exists! Marking as completed since force = FALSE!, 0.001 mins elapsed.
## (s2UI : 3 of 6) Checking if completed file exists!
## 2021-01-28 22:54:29 : (s2UI : 3 of 6) Arrow Exists! Marking as completed since force = FALSE!, 0.001 mins elapsed.
## (s3UI : 4 of 6) Checking if completed file exists!
## 2021-01-28 22:54:29 : (s3UI : 4 of 6) Arrow Exists! Marking as completed since force = FALSE!, 0.002 mins elapsed.
## (s2Inj : 5 of 6) Checking if completed file exists!
## 2021-01-28 22:54:29 : (s2Inj : 5 of 6) Arrow Exists! Marking as completed since force = FALSE!, 0.002 mins elapsed.
## (s1UI : 6 of 6) Checking if completed file exists!
## 2021-01-28 22:54:29 : (s1UI : 6 of 6) Arrow Exists! Marking as completed since force = FALSE!, 0.002 mins elapsed.
## ArchR logging successful to : ../data/scATAC/arrowLogFile.txt
## Using GeneAnnotation set by addArchRGenome(Mm10)!
## Using GeneAnnotation set by addArchRGenome(Mm10)!
## Validating Arrows...
## Getting SampleNames...
## 1 2 3 4 5 6 
## Copying ArrowFiles to Ouptut Directory! If you want to save disk space set copyArrows = FALSE
## 1 2 3 4 5 6 
## Getting Cell Metadata...
## 1 2 3 4 5 6 
## Merging Cell Metadata...
## Initializing ArchRProject...
## 
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##           /   \     |   _  \      /      ||  |  |  | |   _  \     
##          /  ^  \    |  |_)  |    |  ,----'|  |__|  | |  |_)  |    
##         /  /_\  \   |      /     |  |     |   __   | |      /     
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##       /__/     \__\ | _| `._____| \______||__|  |__| | _| `._____|
##     
## ArchR logging to : ArchRLogs/ArchR-addDoubletScores-1f667e7e71cb-Date-2021-01-28_Time-22-54-35.log
## If there is an issue, please report to github with logFile!
## 2021-01-28 22:54:35 : Batch Execution w/ safelapply!, 0 mins elapsed.
## Filtering 1 dims correlated > 0.75 to log10(depth + 1)
## s1Inj (1 of 6) : UMAP Projection R^2 = 0.95591
## s1Inj (1 of 6) : UMAP Projection R^2 = 0.95591

## Filtering 1 dims correlated > 0.75 to log10(depth + 1)
## s3Inj (2 of 6) : UMAP Projection R^2 = 0.97708
## s3Inj (2 of 6) : UMAP Projection R^2 = 0.97708
## Filtering 1 dims correlated > 0.75 to log10(depth + 1)
## s3UI (3 of 6) : UMAP Projection R^2 = 0.98534
## s3UI (3 of 6) : UMAP Projection R^2 = 0.98534
## Filtering 1 dims correlated > 0.75 to log10(depth + 1)
## s2UI (4 of 6) : UMAP Projection R^2 = 0.93469
## s2UI (4 of 6) : UMAP Projection R^2 = 0.93469
## s2Inj (5 of 6) : UMAP Projection R^2 = 0.91338
## s2Inj (5 of 6) : UMAP Projection R^2 = 0.91338
## s1UI (6 of 6) : UMAP Projection R^2 = 0.77624
## s1UI (6 of 6) : UMAP Projection R^2 = 0.77624
## s1UI (6 of 6) : Correlation of UMAP Projection is below 0.9 (normally this is ~0.99)
## This means there is little heterogeneity in your sample and thus doubletCalling is inaccurate.
## force = FALSE, thus returning -1 doubletScores and doubletEnrichments!
## Set force = TRUE if you want to contniue (not recommended).
## s1UI (6 of 6) : Correlation of UMAP Projection is below 0.9 (normally this is ~0.99)
## This means there is little heterogeneity in your sample and thus doubletCalling is inaccurate.
## force = FALSE, thus returning -1 doubletScores and doubletEnrichments!
## Set force = TRUE if you want to contniue (not recommended).
## ArchR logging successful to : ArchRLogs/ArchR-addDoubletScores-1f667e7e71cb-Date-2021-01-28_Time-22-54-35.log
## Checking Inputs...
## ArchR logging to : ArchRLogs/ArchR-addIterativeLSI-1f66581167c0-Date-2021-01-28_Time-23-00-22.log
## If there is an issue, please report to github with logFile!
## 2021-01-28 23:00:22 : Computing Total Accessibility Across All Features, 0.001 mins elapsed.
## 2021-01-28 23:00:24 : Computing Top Features, 0.037 mins elapsed.
## ###########
## 2021-01-28 23:00:24 : Running LSI (1 of 2) on Top Features, 0.041 mins elapsed.
## ###########
## 2021-01-28 23:00:24 : Creating Partial Matrix, 0.042 mins elapsed.
## 2021-01-28 23:00:45 : Computing LSI, 0.382 mins elapsed.
## 2021-01-28 23:01:00 : Identifying Clusters, 0.637 mins elapsed.
## 2021-01-28 23:01:08 : Identified 5 Clusters, 0.77 mins elapsed.
## 2021-01-28 23:01:08 : Saving LSI Iteration, 0.771 mins elapsed.
## 2021-01-28 23:01:22 : Creating Cluster Matrix on the total Group Features, 0.992 mins elapsed.
## 2021-01-28 23:01:58 : Computing Variable Features, 1.607 mins elapsed.
## ###########
## 2021-01-28 23:01:58 : Running LSI (2 of 2) on Variable Features, 1.608 mins elapsed.
## ###########
## 2021-01-28 23:01:58 : Creating Partial Matrix, 1.608 mins elapsed.
## 2021-01-28 23:02:11 : Computing LSI, 1.825 mins elapsed.
## 2021-01-28 23:02:21 : Finished Running IterativeLSI, 1.985 mins elapsed.
## ArchR logging to : ArchRLogs/ArchR-addClusters-1f6645415f5f-Date-2021-01-28_Time-23-02-24.log
## If there is an issue, please report to github with logFile!
## plots: batch correction
p3 <- plotEmbedding(ArchRProj = proj, colorBy = "cellColData", name = "Sample", embedding = "UMAP_cor")
## ArchR logging to : ArchRLogs/ArchR-plotEmbedding-1f66873f839-Date-2021-01-28_Time-23-02-43.log
## If there is an issue, please report to github with logFile!
## Getting UMAP Embedding
## ColorBy = cellColData
## Plotting Embedding
## 1 
## ArchR logging successful to : ArchRLogs/ArchR-plotEmbedding-1f66873f839-Date-2021-01-28_Time-23-02-43.log
p4 <- plotEmbedding(ArchRProj = proj, colorBy = "cellColData", name = "clusters_cor", embedding = "UMAP_cor")
## ArchR logging to : ArchRLogs/ArchR-plotEmbedding-1f6675ad6a84-Date-2021-01-28_Time-23-02-43.log
## If there is an issue, please report to github with logFile!
## Getting UMAP Embedding
## ColorBy = cellColData
## Plotting Embedding
## 1 
## ArchR logging successful to : ArchRLogs/ArchR-plotEmbedding-1f6675ad6a84-Date-2021-01-28_Time-23-02-43.log
ggAlignPlots(p3, p4, type = "h")

Distribution of samples within clusters, and marker genes.

## Getting ImputeWeights
## No imputeWeights found, returning NULL
## ArchR logging to : ArchRLogs/ArchR-plotEmbedding-1f666c35cfb2-Date-2021-01-28_Time-23-02-45.log
## If there is an issue, please report to github with logFile!
## Getting UMAP Embedding
## ColorBy = GeneScoreMatrix
## Getting Matrix Values...
## 2021-01-28 23:02:46 :
## 1 2 3 4 5 6 
## Plotting Embedding
## 1 2 3 4 5 6 7 8 9 10 
## ArchR logging successful to : ArchRLogs/ArchR-plotEmbedding-1f666c35cfb2-Date-2021-01-28_Time-23-02-45.log
## $OMP

## 
## $ASCL1

## 
## $NEUROD1

## 
## $KRT5

## 
## $KRT14

## 
## $TRP63

## 
## $KRTDAP

## 
## $SPRR1A

## 
## $CYP2G1

## 
## $CYP1A2

DA analysis on gene scores, comparing injured vs non-injured HBCs

## ArchR logging to : ArchRLogs/ArchR-plotEmbedding-1f665cbeb053-Date-2021-01-28_Time-23-02-56.log
## If there is an issue, please report to github with logFile!
## Getting UMAP Embedding
## ColorBy = cellColData
## Plotting Embedding
## 1 
## ArchR logging successful to : ArchRLogs/ArchR-plotEmbedding-1f665cbeb053-Date-2021-01-28_Time-23-02-56.log

Plots for paper on full dataset

library(ggplot2)
plotEmbedding(ArchRProj = proj, 
              colorBy = "cellColData", 
              name = "clMerged", 
              embedding = "UMAP_cor")
## ArchR logging to : ArchRLogs/ArchR-plotEmbedding-1f66250bdbb1-Date-2021-01-28_Time-23-02-57.log
## If there is an issue, please report to github with logFile!
## Getting UMAP Embedding
## ColorBy = cellColData
## Plotting Embedding
## 1 
## ArchR logging successful to : ArchRLogs/ArchR-plotEmbedding-1f66250bdbb1-Date-2021-01-28_Time-23-02-57.log

umapDims <- ArchR::getEmbedding(proj, "UMAP_cor")
colnames(umapDims) <- c("UMAP1", "UMAP2")
umapDims$clMerged <- getCellColData(proj, select = "clMerged")$clMerged
umapDims$clMerged[umapDims$clMerged == "Neur"] <- "Neuronal"
umapDims$clMerged[umapDims$clMerged == "Sus"] <- "Sustentacular"

pCellType <- ggplot(umapDims, aes(x=UMAP1, y=UMAP2, color=clMerged)) +
  geom_point(size = 1) + 
  theme_classic() +
  labs(color = "Cell type")

pCellType

Preliminary HBC analysis without integration

## select cells for DA
nCells(proj)
## [1] 4732
proj <- proj[which(clMerged == "HBC"),]
proj <- addUMAP(ArchRProj = proj, reducedDims = "Harmony", force=TRUE, name="UMAP_cor",
  verbose = FALSE)
plotEmbedding(ArchRProj = proj, colorBy = "cellColData", name = "clMerged", embedding = "UMAP_cor")
## ArchR logging to : ArchRLogs/ArchR-plotEmbedding-1f666eaec73e-Date-2021-01-28_Time-23-03-09.log
## If there is an issue, please report to github with logFile!
## Getting UMAP Embedding
## ColorBy = cellColData
## Plotting Embedding
## 1 
## ArchR logging successful to : ArchRLogs/ArchR-plotEmbedding-1f666eaec73e-Date-2021-01-28_Time-23-03-09.log

hbcInj <- vector(length=nCells(proj))
hbcInj[proj@cellColData$treat == "Inj"] <- "hbc_inj"
hbcInj[proj@cellColData$treat == "UI"] <- "hbc_uninj"
proj@cellColData$hbcInj <- hbcInj
plotEmbedding(ArchRProj = proj, colorBy = "cellColData", name = "hbcInj", embedding = "UMAP_cor")
## ArchR logging to : ArchRLogs/ArchR-plotEmbedding-1f66149af037-Date-2021-01-28_Time-23-03-09.log
## If there is an issue, please report to github with logFile!
## Getting UMAP Embedding
## ColorBy = cellColData
## Plotting Embedding
## 1 
## ArchR logging successful to : ArchRLogs/ArchR-plotEmbedding-1f66149af037-Date-2021-01-28_Time-23-03-09.log

markerRes <- getMarkerFeatures(
    ArchRProj = proj, 
    useMatrix = "GeneScoreMatrix", 
    groupBy = "hbcInj",
    bias = c("TSSEnrichment", "log10(nFrags)"),
    testMethod = "wilcoxon",
  verbose = FALSE)
## ArchR logging to : ArchRLogs/ArchR-getMarkerFeatures-1f664810b572-Date-2021-01-28_Time-23-03-10.log
## If there is an issue, please report to github with logFile!
## MatrixClass = Sparse.Double.Matrix
## ArchR logging successful to : ArchRLogs/ArchR-getMarkerFeatures-1f664810b572-Date-2021-01-28_Time-23-03-10.log
markerList <- getMarkers(markerRes, cutOff = "FDR <= 0.01 & Log2FC >= 1.25")
heatmapMarkers <- plotMarkerHeatmap(
  seMarker = markerRes, 
  labelMarkers = markers,
  cutOff = "FDR <= 0.01 & Log2FC >= 1.25", 
  transpose = TRUE
)
## ArchR logging to : ArchRLogs/ArchR-plotMarkerHeatmap-1f6640f87454-Date-2021-01-28_Time-23-03-49.log
## If there is an issue, please report to github with logFile!
## Printing Top Marker Genes:
## hbc_inj:
##  Prss39, Cfc1, Cnga3, Lyg1, Cxcr2, Dnajb3, Ugt1a1, Mir6345, Serpinb2, Htr5b, Il24, Il20, Scarna3a, 4930562F07Rik, Cd48, Ifi211, Mndal, Ifi205, Olfr433, Olfr432
## hbc_uninj:
##  Jsrp1, Msrb3, Fscn2, Lrrc18, Gm16835, Fam135b, Tectb, Gbp7, Oas1c, Cox8b, Ces1b, Mir328, Prss39, Cfc1, Cnga3, Lyg1, Cxcr2, Dnajb3, Ugt1a1, Mir6345
## Identified 212 markers!
## Adding Annotations..
## ArchR logging successful to : ArchRLogs/ArchR-plotMarkerHeatmap-1f6640f87454-Date-2021-01-28_Time-23-03-49.log
# ComplexHeatmap::draw(heatmapMarkers, heatmap_legend_side = "bot", annotation_legend_side = "bot")

Manually merge HBC clusters

plotEmbedding(ArchRProj = proj, colorBy = "cellColData", name = "clusters_cor", embedding = "UMAP_cor")
## ArchR logging to : ArchRLogs/ArchR-plotEmbedding-1f6659e69d65-Date-2021-01-28_Time-23-03-50.log
## If there is an issue, please report to github with logFile!
## Getting UMAP Embedding
## ColorBy = cellColData
## Plotting Embedding
## 1 
## ArchR logging successful to : ArchRLogs/ArchR-plotEmbedding-1f6659e69d65-Date-2021-01-28_Time-23-03-50.log

plotEmbedding(ArchRProj = proj, colorBy = "cellColData", name = "hbcInj", embedding = "UMAP_cor")
## ArchR logging to : ArchRLogs/ArchR-plotEmbedding-1f6633677fff-Date-2021-01-28_Time-23-03-51.log
## If there is an issue, please report to github with logFile!
## Getting UMAP Embedding
## ColorBy = cellColData
## Plotting Embedding
## 1 
## ArchR logging successful to : ArchRLogs/ArchR-plotEmbedding-1f6633677fff-Date-2021-01-28_Time-23-03-51.log

clHBCMerged <- as.character(proj@cellColData$clusters_cor)
clHBCMerged[clHBCMerged %in% c("C3", "C4")] <- "C1_Inj"
clHBCMerged[clHBCMerged %in% c("C9", "C10")] <- "C2_Hybrid"
clHBCMerged[clHBCMerged %in% c("C6", "C7", "C8")] <- "C3_UI"
proj@cellColData$clHBCMerged <- clHBCMerged
plotEmbedding(ArchRProj = proj, colorBy = "cellColData", name = "clHBCMerged", embedding = "UMAP_cor")
## ArchR logging to : ArchRLogs/ArchR-plotEmbedding-1f66500458b6-Date-2021-01-28_Time-23-03-53.log
## If there is an issue, please report to github with logFile!
## Getting UMAP Embedding
## ColorBy = cellColData
## Plotting Embedding
## 1 
## ArchR logging successful to : ArchRLogs/ArchR-plotEmbedding-1f66500458b6-Date-2021-01-28_Time-23-03-53.log

df <- data.frame(cl=proj$clHBCMerged,
                 treatment=proj$treat)

pBar <- ggplot(df, aes(x=treatment, fill=treatment)) + 
  geom_bar(stat="count") +
  facet_wrap(.~cl)
pBar

markerHBCCl <- getMarkerFeatures(
    ArchRProj = proj, 
    useMatrix = "GeneScoreMatrix", 
    groupBy = "clHBCMerged",
    bias = c("TSSEnrichment", "log10(nFrags)"),
    testMethod = "wilcoxon",
  verbose = FALSE)
## ArchR logging to : ArchRLogs/ArchR-getMarkerFeatures-1f664d5c3dc4-Date-2021-01-28_Time-23-03-54.log
## If there is an issue, please report to github with logFile!
## MatrixClass = Sparse.Double.Matrix
## ArchR logging successful to : ArchRLogs/ArchR-getMarkerFeatures-1f664d5c3dc4-Date-2021-01-28_Time-23-03-54.log
markerList_HBCCl <- getMarkers(markerHBCCl, cutOff = "FDR <= 0.01 & Log2FC >= 1.25")
# saveRDS(markerList_HBCCl, file="../data/markerList_atac_ArchR_hbcClusters.rds")

heatmapMarkers <- plotMarkerHeatmap(
  seMarker = markerHBCCl, 
  labelMarkers = markers,
  cutOff = "FDR <= 0.01 & Log2FC >= 1.25", 
  transpose = TRUE
)
## ArchR logging to : ArchRLogs/ArchR-plotMarkerHeatmap-1f6662db3689-Date-2021-01-28_Time-23-04-53.log
## If there is an issue, please report to github with logFile!
## Printing Top Marker Genes:
## C1_Inj:
##  Gm17644, Prss39, Lyg1, Nms, Il1rl1, Slc9a4, Tmem182, Vwc2l, Cxcr2, Ngef, Ugt1a5, Dnajb3, Ugt1a1, Serpinb2, Serpinb10, Il24, Il19, Ifi204, Ifi205, Olfr432
## C2_Hybrid:
##  Dnah7b, Crygf, Fn1, Pax3, Serpinb13, Serpinb3c, Serpinb7, C4bp-ps1, Pigr, Chil1, F5, Gm16548, Olfml2b, Tstd1, Mir194-1, Mir215, 1700007P06Rik, Il22ra2, Rspo3, Gm12070
## C3_UI:
##  Gm8580, Gm4792, Ikzf4, Irf1, Krtap9-5, Tmem235, Fscn2, Mrs2, Mss51, Wdfy4, Lrrc18, Mir5113, Gm5416, 2010005H15Rik, Mir5107, Sec31b, 1700080N15Rik, Gbp7, BC039966, Samd11
## Identified 504 markers!
## Adding Annotations..
## ArchR logging successful to : ArchRLogs/ArchR-plotMarkerHeatmap-1f6662db3689-Date-2021-01-28_Time-23-04-53.log
heatmapHBC <- plotMarkerHeatmap(
  seMarker = markerHBCCl, 
  cutOff = "FDR <= 0.01 & Log2FC >= 1.25", 
  transpose = TRUE
)
## ArchR logging to : ArchRLogs/ArchR-plotMarkerHeatmap-1f6625d98f13-Date-2021-01-28_Time-23-04-53.log
## If there is an issue, please report to github with logFile!
## Printing Top Marker Genes:
## C1_Inj:
##  Gm17644, Prss39, Lyg1, Nms, Il1rl1, Slc9a4, Tmem182, Vwc2l, Cxcr2, Ngef, Ugt1a5, Dnajb3, Ugt1a1, Serpinb2, Serpinb10, Il24, Il19, Ifi204, Ifi205, Olfr432
## C2_Hybrid:
##  Dnah7b, Crygf, Fn1, Pax3, Serpinb13, Serpinb3c, Serpinb7, C4bp-ps1, Pigr, Chil1, F5, Gm16548, Olfml2b, Tstd1, Mir194-1, Mir215, 1700007P06Rik, Il22ra2, Rspo3, Gm12070
## C3_UI:
##  Gm8580, Gm4792, Ikzf4, Irf1, Krtap9-5, Tmem235, Fscn2, Mrs2, Mss51, Wdfy4, Lrrc18, Mir5113, Gm5416, 2010005H15Rik, Mir5107, Sec31b, 1700080N15Rik, Gbp7, BC039966, Samd11
## Identified 504 markers!
## Preparing Main Heatmap..
## ArchR logging successful to : ArchRLogs/ArchR-plotMarkerHeatmap-1f6625d98f13-Date-2021-01-28_Time-23-04-53.log
ComplexHeatmap::draw(heatmapHBC, heatmap_legend_side = "bot", annotation_legend_side = "bot")

Plot for paper on preliminary HBC analysis

library(ggplot2)
umapDimsHBC <- ArchR::getEmbedding(proj, "UMAP_cor")
colnames(umapDimsHBC) <- c("UMAP1", "UMAP2")
umapDimsHBC$clHBCMerged <- getCellColData(proj, select = "clHBCMerged")$clHBCMerged
umapDimsHBC$treat <- getCellColData(proj, select = "treat")$treat

# cell state
pstate <- ggplot(umapDimsHBC, aes(x=UMAP1, y=UMAP2, color=clHBCMerged)) +
  geom_point(size = 1) + 
  theme_classic() +
  labs(color = "Cell state")

# injury treatment
pinj <- ggplot(umapDimsHBC, aes(x=UMAP1, y=UMAP2, color=treat)) +
  geom_point(size = 1) + 
  theme_classic() +
  labs(color = "Treatment") + 
  scale_color_manual(values = c("orange", "darkseagreen3")) 

cowplot::plot_grid(pstate, pinj, nrow=1)

ggsave("../plots/hbcDR.png", width=10)
## Saving 10 x 5 in image

Export gene activity data for custom downstream analysis

useMatrix="GeneScoreMatrix"
ArrowFiles <- getArrowFiles(proj)
featureDF <- ArchR:::.getFeatureDF(head(ArrowFiles, 2), "GeneScoreMatrix")
geneMat <- ArchR:::.getPartialMatrix(ArrowFiles, 
            featureDF = featureDF, threads = 1, useMatrix = "GeneScoreMatrix", 
            cellNames = rownames(proj@cellColData), progress = FALSE)
rownames(geneMat) <- featureDF$name
saveRDS(geneMat, file="../data/scATAC/geneMatArchrHbc.rds")

Create pseudobulk for peak calling

## ArchR logging to : ArchRLogs/ArchR-addGroupCoverages-1f664c899308-Date-2021-01-28_Time-23-04-58.log
## If there is an issue, please report to github with logFile!
## C1_Inj (1 of 3) : CellGroups N = 4
## C2_Hybrid (2 of 3) : CellGroups N = 6
## C3_UI (3 of 3) : CellGroups N = 5
## 2021-01-28 23:04:59 : Creating Coverage Files!, 0.008 mins elapsed.
## 2021-01-28 23:04:59 : Batch Execution w/ safelapply!, 0.008 mins elapsed.
## 2021-01-28 23:04:59 : Group C1_Inj._.s1Inj (1 of 15) : Creating Group Coverage File : C1_Inj._.s1Inj.insertions.coverage.h5, 0.01 mins elapsed.
## Number of Cells = 500
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:05:19 : Group C1_Inj._.s2Inj (2 of 15) : Creating Group Coverage File : C1_Inj._.s2Inj.insertions.coverage.h5, 0.343 mins elapsed.
## Number of Cells = 263
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:05:37 : Group C1_Inj._.s3Inj (3 of 15) : Creating Group Coverage File : C1_Inj._.s3Inj.insertions.coverage.h5, 0.653 mins elapsed.
## Number of Cells = 164
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:05:55 : Group C1_Inj._.Other (4 of 15) : Creating Group Coverage File : C1_Inj._.Other.insertions.coverage.h5, 0.937 mins elapsed.
## Number of Cells = 54
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:06:10 : Group C2_Hybrid._.s3UI (5 of 15) : Creating Group Coverage File : C2_Hybrid._.s3UI.insertions.coverage.h5, 1.202 mins elapsed.
## Number of Cells = 377
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:06:32 : Group C2_Hybrid._.s3Inj (6 of 15) : Creating Group Coverage File : C2_Hybrid._.s3Inj.insertions.coverage.h5, 1.568 mins elapsed.
## Number of Cells = 346
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:06:51 : Group C2_Hybrid._.s1Inj (7 of 15) : Creating Group Coverage File : C2_Hybrid._.s1Inj.insertions.coverage.h5, 1.882 mins elapsed.
## Number of Cells = 287
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:07:08 : Group C2_Hybrid._.s2UI (8 of 15) : Creating Group Coverage File : C2_Hybrid._.s2UI.insertions.coverage.h5, 2.16 mins elapsed.
## Number of Cells = 160
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:07:23 : Group C2_Hybrid._.s2Inj (9 of 15) : Creating Group Coverage File : C2_Hybrid._.s2Inj.insertions.coverage.h5, 2.406 mins elapsed.
## Number of Cells = 93
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:07:37 : Group C2_Hybrid._.s1UI (10 of 15) : Creating Group Coverage File : C2_Hybrid._.s1UI.insertions.coverage.h5, 2.651 mins elapsed.
## Number of Cells = 87
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:07:51 : Group C3_UI._.s2UI (11 of 15) : Creating Group Coverage File : C3_UI._.s2UI.insertions.coverage.h5, 2.884 mins elapsed.
## Number of Cells = 500
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:08:10 : Group C3_UI._.s1UI (12 of 15) : Creating Group Coverage File : C3_UI._.s1UI.insertions.coverage.h5, 3.203 mins elapsed.
## Number of Cells = 355
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:08:27 : Group C3_UI._.s3UI (13 of 15) : Creating Group Coverage File : C3_UI._.s3UI.insertions.coverage.h5, 3.484 mins elapsed.
## Number of Cells = 178
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:08:43 : Group C3_UI._.s3Inj (14 of 15) : Creating Group Coverage File : C3_UI._.s3Inj.insertions.coverage.h5, 3.742 mins elapsed.
## Number of Cells = 55
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:08:57 : Group C3_UI._.Other (15 of 15) : Creating Group Coverage File : C3_UI._.Other.insertions.coverage.h5, 3.972 mins elapsed.
## Number of Cells = 51
## Coverage File Exists!
## Added Coverage Group
## Added Metadata Group
## Added ArrowCoverage Class
## Added Coverage/Info
## Added Coverage/Info/CellNames
## 2021-01-28 23:09:12 : Adding Kmer Bias to Coverage Files!, 4.223 mins elapsed.
## Kmer Bias chr1 (1 of 21)
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## Coverage File chr6 (7 of 15)
## Coverage File chr6 (8 of 15)
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## Coverage File chr7 (6 of 15)
## Coverage File chr7 (7 of 15)
## Coverage File chr7 (8 of 15)
## Coverage File chr7 (9 of 15)
## Coverage File chr7 (10 of 15)
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## Coverage File chr7 (15 of 15)
## Kmer Bias chr8 (18 of 21)
## chr8 Coverage File chr8 (1 of 15)
## Coverage File chr8 (2 of 15)
## Coverage File chr8 (3 of 15)
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## Coverage File chr8 (5 of 15)
## Coverage File chr8 (6 of 15)
## Coverage File chr8 (7 of 15)
## Coverage File chr8 (8 of 15)
## Coverage File chr8 (9 of 15)
## Coverage File chr8 (10 of 15)
## Coverage File chr8 (11 of 15)
## Coverage File chr8 (12 of 15)
## Coverage File chr8 (13 of 15)
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## Coverage File chr8 (15 of 15)
## Kmer Bias chr9 (19 of 21)
## chr9 Coverage File chr9 (1 of 15)
## Coverage File chr9 (2 of 15)
## Coverage File chr9 (3 of 15)
## Coverage File chr9 (4 of 15)
## Coverage File chr9 (5 of 15)
## Coverage File chr9 (6 of 15)
## Coverage File chr9 (7 of 15)
## Coverage File chr9 (8 of 15)
## Coverage File chr9 (9 of 15)
## Coverage File chr9 (10 of 15)
## Coverage File chr9 (11 of 15)
## Coverage File chr9 (12 of 15)
## Coverage File chr9 (13 of 15)
## Coverage File chr9 (14 of 15)
## Coverage File chr9 (15 of 15)
## Kmer Bias chrX (20 of 21)
## chrX Coverage File chrX (1 of 15)
## Coverage File chrX (2 of 15)
## Coverage File chrX (3 of 15)
## Coverage File chrX (4 of 15)
## Coverage File chrX (5 of 15)
## Coverage File chrX (6 of 15)
## Coverage File chrX (7 of 15)
## Coverage File chrX (8 of 15)
## Coverage File chrX (9 of 15)
## Coverage File chrX (10 of 15)
## Coverage File chrX (11 of 15)
## Coverage File chrX (12 of 15)
## Coverage File chrX (13 of 15)
## Coverage File chrX (14 of 15)
## Coverage File chrX (15 of 15)
## Kmer Bias chrY (21 of 21)
## chrY Coverage File chrY (1 of 15)
## Coverage File chrY (2 of 15)
## Coverage File chrY (3 of 15)
## Coverage File chrY (4 of 15)
## Coverage File chrY (5 of 15)
## Coverage File chrY (6 of 15)
## Coverage File chrY (7 of 15)
## Coverage File chrY (8 of 15)
## Coverage File chrY (9 of 15)
## Coverage File chrY (10 of 15)
## Coverage File chrY (11 of 15)
## Coverage File chrY (12 of 15)
## Coverage File chrY (13 of 15)
## Coverage File chrY (14 of 15)
## Coverage File chrY (15 of 15)
## Completed Kmer Bias Calculation
## Adding Kmer Bias (1 of 15)
## Adding Kmer Bias (2 of 15)
## Adding Kmer Bias (3 of 15)
## Adding Kmer Bias (4 of 15)
## Adding Kmer Bias (5 of 15)
## Adding Kmer Bias (6 of 15)
## Adding Kmer Bias (7 of 15)
## Adding Kmer Bias (8 of 15)
## Adding Kmer Bias (9 of 15)
## Adding Kmer Bias (10 of 15)
## Adding Kmer Bias (11 of 15)
## Adding Kmer Bias (12 of 15)
## Adding Kmer Bias (13 of 15)
## Adding Kmer Bias (14 of 15)
## Adding Kmer Bias (15 of 15)
## 2021-01-28 23:13:57 : Finished Creation of Coverage Files!, 8.979 mins elapsed.
## ArchR logging successful to : ArchRLogs/ArchR-addGroupCoverages-1f664c899308-Date-2021-01-28_Time-23-04-58.log

Peak calling

## ArchR logging to : ArchRLogs/ArchR-addReproduciblePeakSet-1f662b948629-Date-2021-01-28_Time-23-13-58.log
## If there is an issue, please report to github with logFile!
## Calling Peaks with Macs2
## 2021-01-28 23:13:58 : Peak Calling Parameters!, 0.004 mins elapsed.
##               Group nCells nCellsUsed nReplicates nMin nMax maxPeaks
## C1_Inj       C1_Inj   1486        981           4   54  500   150000
## C2_Hybrid C2_Hybrid   1350       1350           6   87  377   150000
## C3_UI         C3_UI   1240       1139           5   51  500   150000
## 2021-01-28 23:13:58 : Batching Peak Calls!, 0.004 mins elapsed.
## 2021-01-28 23:13:58 : Batch Execution w/ safelapply!, 0 mins elapsed.
## 2021-01-28 23:13:58 : Group 1 of 15, Calling Peaks with MACS2!, 0 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C1_Inj._.s1Inj-1 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C1_Inj._.s1Inj-1.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:15:30 : Group 2 of 15, Calling Peaks with MACS2!, 1.53 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C1_Inj._.s2Inj-2 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C1_Inj._.s2Inj-2.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:16:52 : Group 3 of 15, Calling Peaks with MACS2!, 2.909 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C1_Inj._.s3Inj-3 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C1_Inj._.s3Inj-3.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:18:07 : Group 4 of 15, Calling Peaks with MACS2!, 4.16 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C1_Inj._.Other-4 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C1_Inj._.Other-4.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:19:02 : Group 5 of 15, Calling Peaks with MACS2!, 5.067 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C2_Hybrid._.s3UI-5 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C2_Hybrid._.s3UI-5.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:20:44 : Group 6 of 15, Calling Peaks with MACS2!, 6.77 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C2_Hybrid._.s3Inj-6 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C2_Hybrid._.s3Inj-6.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:22:08 : Group 7 of 15, Calling Peaks with MACS2!, 8.163 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C2_Hybrid._.s1Inj-7 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C2_Hybrid._.s1Inj-7.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:23:12 : Group 8 of 15, Calling Peaks with MACS2!, 9.236 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C2_Hybrid._.s2UI-8 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C2_Hybrid._.s2UI-8.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:24:01 : Group 9 of 15, Calling Peaks with MACS2!, 10.06 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C2_Hybrid._.s2Inj-9 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C2_Hybrid._.s2Inj-9.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:24:54 : Group 10 of 15, Calling Peaks with MACS2!, 10.94 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C2_Hybrid._.s1UI-10 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C2_Hybrid._.s1UI-10.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:25:29 : Group 11 of 15, Calling Peaks with MACS2!, 11.523 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C3_UI._.s2UI-11 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C3_UI._.s2UI-11.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:26:45 : Group 12 of 15, Calling Peaks with MACS2!, 12.79 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C3_UI._.s1UI-12 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C3_UI._.s1UI-12.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:27:53 : Group 13 of 15, Calling Peaks with MACS2!, 13.928 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C3_UI._.s3UI-13 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C3_UI._.s3UI-13.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:28:48 : Group 14 of 15, Calling Peaks with MACS2!, 14.834 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C3_UI._.s3Inj-14 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C3_UI._.s3Inj-14.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:29:26 : Group 15 of 15, Calling Peaks with MACS2!, 15.468 mins elapsed.
## Running Macs2 with Params : macs2 callpeak -g 1.87e+09 --name C3_UI._.Other-15 --treatment /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds/C3_UI._.Other-15.insertions.bed --outdir /Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/InsertionBeds --format BED --call-summits --keep-dup all --nomodel --nolambda --shift -75 --extsize 150 -q 0.1
## 2021-01-28 23:29:56 : Identifying Reproducible Peaks!, 15.979 mins elapsed.
## Annotating Peaks : Nearest Gene
## Annotating Peaks : Gene
## Annotating Peaks : TSS
## Annotating Peaks : GC
## [1] "/Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/C1_Inj-reproduciblePeaks.gr.rds"
## Annotating Peaks : Nearest Gene
## Annotating Peaks : Gene
## Annotating Peaks : TSS
## Annotating Peaks : GC
## [1] "/Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/C2_Hybrid-reproduciblePeaks.gr.rds"
## Annotating Peaks : Nearest Gene
## Annotating Peaks : Gene
## Annotating Peaks : TSS
## Annotating Peaks : GC
## [1] "/Users/koenvandenberge/Dropbox/research/brainStat/hbcRegenGithub/data/scATAC/archR_samples_pairwise/PeakCalls/C3_UI-reproduciblePeaks.gr.rds"
## 2021-01-28 23:30:29 : Creating Union Peak Set!, 16.528 mins elapsed.
## Converged after 7 iterations!
## Plotting Ggplot!
## 2021-01-28 23:30:33 : Finished Creating Union Peak Set (183712)!, 16.595 mins elapsed.
## ArchR logging to : ArchRLogs/ArchR-addPeakMatrix-1f66693917bf-Date-2021-01-28_Time-23-30-33.log
## If there is an issue, please report to github with logFile!
## 2021-01-28 23:30:33 : Batch Execution w/ safelapply!, 0 mins elapsed.
## 2021-01-28 23:30:34 : Adding s1Inj to PeakMatrix for Chr (1 of 20)!, 0.009 mins elapsed.
## 2021-01-28 23:30:36 : Adding s1Inj to PeakMatrix for Chr (2 of 20)!, 0.042 mins elapsed.
## 2021-01-28 23:30:38 : Adding s1Inj to PeakMatrix for Chr (3 of 20)!, 0.074 mins elapsed.
## 2021-01-28 23:30:40 : Adding s1Inj to PeakMatrix for Chr (4 of 20)!, 0.104 mins elapsed.
## 2021-01-28 23:30:42 : Adding s1Inj to PeakMatrix for Chr (5 of 20)!, 0.135 mins elapsed.
## 2021-01-28 23:30:43 : Adding s1Inj to PeakMatrix for Chr (6 of 20)!, 0.165 mins elapsed.
## 2021-01-28 23:30:45 : Adding s1Inj to PeakMatrix for Chr (7 of 20)!, 0.195 mins elapsed.
## 2021-01-28 23:30:47 : Adding s1Inj to PeakMatrix for Chr (8 of 20)!, 0.226 mins elapsed.
## 2021-01-28 23:30:49 : Adding s1Inj to PeakMatrix for Chr (9 of 20)!, 0.256 mins elapsed.
## 2021-01-28 23:30:50 : Adding s1Inj to PeakMatrix for Chr (10 of 20)!, 0.284 mins elapsed.
## 2021-01-28 23:30:52 : Adding s1Inj to PeakMatrix for Chr (11 of 20)!, 0.312 mins elapsed.
## 2021-01-28 23:30:54 : Adding s1Inj to PeakMatrix for Chr (12 of 20)!, 0.344 mins elapsed.
## 2021-01-28 23:30:56 : Adding s1Inj to PeakMatrix for Chr (13 of 20)!, 0.371 mins elapsed.
## 2021-01-28 23:30:57 : Adding s1Inj to PeakMatrix for Chr (14 of 20)!, 0.398 mins elapsed.
## 2021-01-28 23:30:59 : Adding s1Inj to PeakMatrix for Chr (15 of 20)!, 0.425 mins elapsed.
## 2021-01-28 23:31:01 : Adding s1Inj to PeakMatrix for Chr (16 of 20)!, 0.452 mins elapsed.
## 2021-01-28 23:31:02 : Adding s1Inj to PeakMatrix for Chr (17 of 20)!, 0.478 mins elapsed.
## 2021-01-28 23:31:04 : Adding s1Inj to PeakMatrix for Chr (18 of 20)!, 0.506 mins elapsed.
## 2021-01-28 23:31:05 : Adding s1Inj to PeakMatrix for Chr (19 of 20)!, 0.531 mins elapsed.
## 2021-01-28 23:31:07 : Adding s1Inj to PeakMatrix for Chr (20 of 20)!, 0.557 mins elapsed.
## 2021-01-28 23:31:09 : Adding s3Inj to PeakMatrix for Chr (1 of 20)!, 0.009 mins elapsed.
## 2021-01-28 23:31:10 : Adding s3Inj to PeakMatrix for Chr (2 of 20)!, 0.036 mins elapsed.
## 2021-01-28 23:31:12 : Adding s3Inj to PeakMatrix for Chr (3 of 20)!, 0.064 mins elapsed.
## 2021-01-28 23:31:14 : Adding s3Inj to PeakMatrix for Chr (4 of 20)!, 0.09 mins elapsed.
## 2021-01-28 23:31:15 : Adding s3Inj to PeakMatrix for Chr (5 of 20)!, 0.117 mins elapsed.
## 2021-01-28 23:31:17 : Adding s3Inj to PeakMatrix for Chr (6 of 20)!, 0.144 mins elapsed.
## 2021-01-28 23:31:18 : Adding s3Inj to PeakMatrix for Chr (7 of 20)!, 0.17 mins elapsed.
## 2021-01-28 23:31:20 : Adding s3Inj to PeakMatrix for Chr (8 of 20)!, 0.197 mins elapsed.
## 2021-01-28 23:31:22 : Adding s3Inj to PeakMatrix for Chr (9 of 20)!, 0.223 mins elapsed.
## 2021-01-28 23:31:23 : Adding s3Inj to PeakMatrix for Chr (10 of 20)!, 0.248 mins elapsed.
## 2021-01-28 23:31:25 : Adding s3Inj to PeakMatrix for Chr (11 of 20)!, 0.274 mins elapsed.
## 2021-01-28 23:31:26 : Adding s3Inj to PeakMatrix for Chr (12 of 20)!, 0.301 mins elapsed.
## 2021-01-28 23:31:28 : Adding s3Inj to PeakMatrix for Chr (13 of 20)!, 0.326 mins elapsed.
## 2021-01-28 23:31:29 : Adding s3Inj to PeakMatrix for Chr (14 of 20)!, 0.351 mins elapsed.
## 2021-01-28 23:31:31 : Adding s3Inj to PeakMatrix for Chr (15 of 20)!, 0.375 mins elapsed.
## 2021-01-28 23:31:32 : Adding s3Inj to PeakMatrix for Chr (16 of 20)!, 0.399 mins elapsed.
## 2021-01-28 23:31:34 : Adding s3Inj to PeakMatrix for Chr (17 of 20)!, 0.423 mins elapsed.
## 2021-01-28 23:31:35 : Adding s3Inj to PeakMatrix for Chr (18 of 20)!, 0.448 mins elapsed.
## 2021-01-28 23:31:37 : Adding s3Inj to PeakMatrix for Chr (19 of 20)!, 0.471 mins elapsed.
## 2021-01-28 23:31:38 : Adding s3Inj to PeakMatrix for Chr (20 of 20)!, 0.495 mins elapsed.
## 2021-01-28 23:31:40 : Adding s3UI to PeakMatrix for Chr (1 of 20)!, 0.009 mins elapsed.
## 2021-01-28 23:31:42 : Adding s3UI to PeakMatrix for Chr (2 of 20)!, 0.036 mins elapsed.
## 2021-01-28 23:31:43 : Adding s3UI to PeakMatrix for Chr (3 of 20)!, 0.063 mins elapsed.
## 2021-01-28 23:31:45 : Adding s3UI to PeakMatrix for Chr (4 of 20)!, 0.089 mins elapsed.
## 2021-01-28 23:31:46 : Adding s3UI to PeakMatrix for Chr (5 of 20)!, 0.116 mins elapsed.
## 2021-01-28 23:31:48 : Adding s3UI to PeakMatrix for Chr (6 of 20)!, 0.143 mins elapsed.
## 2021-01-28 23:31:50 : Adding s3UI to PeakMatrix for Chr (7 of 20)!, 0.169 mins elapsed.
## 2021-01-28 23:31:51 : Adding s3UI to PeakMatrix for Chr (8 of 20)!, 0.197 mins elapsed.
## 2021-01-28 23:31:53 : Adding s3UI to PeakMatrix for Chr (9 of 20)!, 0.223 mins elapsed.
## 2021-01-28 23:31:54 : Adding s3UI to PeakMatrix for Chr (10 of 20)!, 0.248 mins elapsed.
## 2021-01-28 23:31:56 : Adding s3UI to PeakMatrix for Chr (11 of 20)!, 0.274 mins elapsed.
## 2021-01-28 23:31:57 : Adding s3UI to PeakMatrix for Chr (12 of 20)!, 0.302 mins elapsed.
## 2021-01-28 23:31:59 : Adding s3UI to PeakMatrix for Chr (13 of 20)!, 0.326 mins elapsed.
## 2021-01-28 23:32:00 : Adding s3UI to PeakMatrix for Chr (14 of 20)!, 0.351 mins elapsed.
## 2021-01-28 23:32:02 : Adding s3UI to PeakMatrix for Chr (15 of 20)!, 0.375 mins elapsed.
## 2021-01-28 23:32:03 : Adding s3UI to PeakMatrix for Chr (16 of 20)!, 0.4 mins elapsed.
## 2021-01-28 23:32:05 : Adding s3UI to PeakMatrix for Chr (17 of 20)!, 0.424 mins elapsed.
## 2021-01-28 23:32:06 : Adding s3UI to PeakMatrix for Chr (18 of 20)!, 0.449 mins elapsed.
## 2021-01-28 23:32:08 : Adding s3UI to PeakMatrix for Chr (19 of 20)!, 0.472 mins elapsed.
## 2021-01-28 23:32:09 : Adding s3UI to PeakMatrix for Chr (20 of 20)!, 0.496 mins elapsed.
## 2021-01-28 23:32:11 : Adding s2UI to PeakMatrix for Chr (1 of 20)!, 0.009 mins elapsed.
## 2021-01-28 23:32:13 : Adding s2UI to PeakMatrix for Chr (2 of 20)!, 0.035 mins elapsed.
## 2021-01-28 23:32:14 : Adding s2UI to PeakMatrix for Chr (3 of 20)!, 0.061 mins elapsed.
## 2021-01-28 23:32:16 : Adding s2UI to PeakMatrix for Chr (4 of 20)!, 0.086 mins elapsed.
## 2021-01-28 23:32:17 : Adding s2UI to PeakMatrix for Chr (5 of 20)!, 0.112 mins elapsed.
## 2021-01-28 23:32:19 : Adding s2UI to PeakMatrix for Chr (6 of 20)!, 0.137 mins elapsed.
## 2021-01-28 23:32:20 : Adding s2UI to PeakMatrix for Chr (7 of 20)!, 0.162 mins elapsed.
## 2021-01-28 23:32:22 : Adding s2UI to PeakMatrix for Chr (8 of 20)!, 0.189 mins elapsed.
## 2021-01-28 23:32:23 : Adding s2UI to PeakMatrix for Chr (9 of 20)!, 0.214 mins elapsed.
## 2021-01-28 23:32:25 : Adding s2UI to PeakMatrix for Chr (10 of 20)!, 0.238 mins elapsed.
## 2021-01-28 23:32:26 : Adding s2UI to PeakMatrix for Chr (11 of 20)!, 0.263 mins elapsed.
## 2021-01-28 23:32:28 : Adding s2UI to PeakMatrix for Chr (12 of 20)!, 0.29 mins elapsed.
## 2021-01-28 23:32:29 : Adding s2UI to PeakMatrix for Chr (13 of 20)!, 0.314 mins elapsed.
## 2021-01-28 23:32:31 : Adding s2UI to PeakMatrix for Chr (14 of 20)!, 0.337 mins elapsed.
## 2021-01-28 23:32:32 : Adding s2UI to PeakMatrix for Chr (15 of 20)!, 0.361 mins elapsed.
## 2021-01-28 23:32:34 : Adding s2UI to PeakMatrix for Chr (16 of 20)!, 0.385 mins elapsed.
## 2021-01-28 23:32:35 : Adding s2UI to PeakMatrix for Chr (17 of 20)!, 0.409 mins elapsed.
## 2021-01-28 23:32:37 : Adding s2UI to PeakMatrix for Chr (18 of 20)!, 0.433 mins elapsed.
## 2021-01-28 23:32:38 : Adding s2UI to PeakMatrix for Chr (19 of 20)!, 0.456 mins elapsed.
## 2021-01-28 23:32:39 : Adding s2UI to PeakMatrix for Chr (20 of 20)!, 0.479 mins elapsed.
## 2021-01-28 23:32:41 : Adding s2Inj to PeakMatrix for Chr (1 of 20)!, 0.009 mins elapsed.
## 2021-01-28 23:32:43 : Adding s2Inj to PeakMatrix for Chr (2 of 20)!, 0.035 mins elapsed.
## 2021-01-28 23:32:44 : Adding s2Inj to PeakMatrix for Chr (3 of 20)!, 0.061 mins elapsed.
## 2021-01-28 23:32:46 : Adding s2Inj to PeakMatrix for Chr (4 of 20)!, 0.086 mins elapsed.
## 2021-01-28 23:32:47 : Adding s2Inj to PeakMatrix for Chr (5 of 20)!, 0.111 mins elapsed.
## 2021-01-28 23:32:49 : Adding s2Inj to PeakMatrix for Chr (6 of 20)!, 0.136 mins elapsed.
## 2021-01-28 23:32:50 : Adding s2Inj to PeakMatrix for Chr (7 of 20)!, 0.161 mins elapsed.
## 2021-01-28 23:32:52 : Adding s2Inj to PeakMatrix for Chr (8 of 20)!, 0.186 mins elapsed.
## 2021-01-28 23:32:53 : Adding s2Inj to PeakMatrix for Chr (9 of 20)!, 0.21 mins elapsed.
## 2021-01-28 23:32:55 : Adding s2Inj to PeakMatrix for Chr (10 of 20)!, 0.234 mins elapsed.
## 2021-01-28 23:32:56 : Adding s2Inj to PeakMatrix for Chr (11 of 20)!, 0.258 mins elapsed.
## 2021-01-28 23:32:58 : Adding s2Inj to PeakMatrix for Chr (12 of 20)!, 0.284 mins elapsed.
## 2021-01-28 23:32:59 : Adding s2Inj to PeakMatrix for Chr (13 of 20)!, 0.307 mins elapsed.
## 2021-01-28 23:33:01 : Adding s2Inj to PeakMatrix for Chr (14 of 20)!, 0.331 mins elapsed.
## 2021-01-28 23:33:02 : Adding s2Inj to PeakMatrix for Chr (15 of 20)!, 0.354 mins elapsed.
## 2021-01-28 23:33:03 : Adding s2Inj to PeakMatrix for Chr (16 of 20)!, 0.378 mins elapsed.
## 2021-01-28 23:33:05 : Adding s2Inj to PeakMatrix for Chr (17 of 20)!, 0.4 mins elapsed.
## 2021-01-28 23:33:06 : Adding s2Inj to PeakMatrix for Chr (18 of 20)!, 0.424 mins elapsed.
## 2021-01-28 23:33:07 : Adding s2Inj to PeakMatrix for Chr (19 of 20)!, 0.447 mins elapsed.
## 2021-01-28 23:33:09 : Adding s2Inj to PeakMatrix for Chr (20 of 20)!, 0.47 mins elapsed.
## 2021-01-28 23:33:11 : Adding s1UI to PeakMatrix for Chr (1 of 20)!, 0.009 mins elapsed.
## 2021-01-28 23:33:12 : Adding s1UI to PeakMatrix for Chr (2 of 20)!, 0.034 mins elapsed.
## 2021-01-28 23:33:14 : Adding s1UI to PeakMatrix for Chr (3 of 20)!, 0.059 mins elapsed.
## 2021-01-28 23:33:15 : Adding s1UI to PeakMatrix for Chr (4 of 20)!, 0.081 mins elapsed.
## 2021-01-28 23:33:17 : Adding s1UI to PeakMatrix for Chr (5 of 20)!, 0.105 mins elapsed.
## 2021-01-28 23:33:18 : Adding s1UI to PeakMatrix for Chr (6 of 20)!, 0.129 mins elapsed.
## 2021-01-28 23:33:19 : Adding s1UI to PeakMatrix for Chr (7 of 20)!, 0.153 mins elapsed.
## 2021-01-28 23:33:21 : Adding s1UI to PeakMatrix for Chr (8 of 20)!, 0.178 mins elapsed.
## 2021-01-28 23:33:22 : Adding s1UI to PeakMatrix for Chr (9 of 20)!, 0.202 mins elapsed.
## 2021-01-28 23:33:24 : Adding s1UI to PeakMatrix for Chr (10 of 20)!, 0.225 mins elapsed.
## 2021-01-28 23:33:25 : Adding s1UI to PeakMatrix for Chr (11 of 20)!, 0.249 mins elapsed.
## 2021-01-28 23:33:27 : Adding s1UI to PeakMatrix for Chr (12 of 20)!, 0.274 mins elapsed.
## 2021-01-28 23:33:28 : Adding s1UI to PeakMatrix for Chr (13 of 20)!, 0.297 mins elapsed.
## 2021-01-28 23:33:29 : Adding s1UI to PeakMatrix for Chr (14 of 20)!, 0.32 mins elapsed.
## 2021-01-28 23:33:31 : Adding s1UI to PeakMatrix for Chr (15 of 20)!, 0.343 mins elapsed.
## 2021-01-28 23:33:32 : Adding s1UI to PeakMatrix for Chr (16 of 20)!, 0.366 mins elapsed.
## 2021-01-28 23:33:34 : Adding s1UI to PeakMatrix for Chr (17 of 20)!, 0.388 mins elapsed.
## 2021-01-28 23:33:35 : Adding s1UI to PeakMatrix for Chr (18 of 20)!, 0.412 mins elapsed.
## 2021-01-28 23:33:36 : Adding s1UI to PeakMatrix for Chr (19 of 20)!, 0.434 mins elapsed.
## 2021-01-28 23:33:38 : Adding s1UI to PeakMatrix for Chr (20 of 20)!, 0.456 mins elapsed.
## ArchR logging successful to : ArchRLogs/ArchR-addPeakMatrix-1f66693917bf-Date-2021-01-28_Time-23-30-33.log

Motif enrichment

# add motifs
projPeaks <- addMotifAnnotations(ArchRProj = projPeaks, 
                                motifSet = "cisbp", 
                                name = "Motif",
                                force=TRUE)
## No methods found in package 'IRanges' for request: 'score' when loading 'TFBSTools'
## ArchR logging to : ArchRLogs/ArchR-addMotifAnnotations-1f6655a1dfae-Date-2021-01-28_Time-23-33-39.log
## If there is an issue, please report to github with logFile!
## 2021-01-28 23:33:41 : Gettting Motif Set, Species : Mus musculus, 0.001 mins elapsed.
## Using version 2 motifs!
## 2021-01-28 23:33:43 : Finding Motif Positions with motifmatchr!, 0.03 mins elapsed.
## 2021-01-28 23:36:27 : Creating Motif Overlap Matrix, 2.763 mins elapsed.
## 2021-01-28 23:36:30 : Finished Getting Motif Info!, 2.82 mins elapsed.
## ArchR logging successful to : ArchRLogs/ArchR-addMotifAnnotations-1f6655a1dfae-Date-2021-01-28_Time-23-33-39.log
# get markers
peakMarkersHBC <- getMarkerFeatures(
    ArchRProj = projPeaks, 
    useMatrix = "PeakMatrix", 
    groupBy = "clHBCMerged",
    bias = c("TSSEnrichment", "log10(nFrags)"),
    testMethod = "wilcoxon",
  verbose = FALSE)
## ArchR logging to : ArchRLogs/ArchR-getMarkerFeatures-1f667664446d-Date-2021-01-28_Time-23-36-33.log
## If there is an issue, please report to github with logFile!
## MatrixClass = Sparse.Integer.Matrix
## ArchR logging successful to : ArchRLogs/ArchR-getMarkerFeatures-1f667664446d-Date-2021-01-28_Time-23-36-33.log
peakMarkerListHBC <- getMarkers(peakMarkersHBC, cutOff = "FDR <= 0.01 & Log2FC >= 1.25")
saveRDS(markerList_HBCCl, file="../data/scATAC/markerList_atac_ArchR_hbcClusters.rds")

heatmapMarkers <- plotMarkerHeatmap(
  seMarker = peakMarkersHBC, 
  cutOff = "FDR <= 0.01 & Log2FC >= 1.25", 
  transpose = TRUE
)
## ArchR logging to : ArchRLogs/ArchR-plotMarkerHeatmap-1f6630c88cd4-Date-2021-01-28_Time-23-37-43.log
## If there is an issue, please report to github with logFile!
## Identified 24150 markers!
## Preparing Main Heatmap..
## ArchR logging successful to : ArchRLogs/ArchR-plotMarkerHeatmap-1f6630c88cd4-Date-2021-01-28_Time-23-37-43.log
ComplexHeatmap::draw(heatmapMarkers, heatmap_legend_side = "bot", annotation_legend_side = "bot")


## upregulated motifs 
motifsUp <- peakAnnoEnrichment(
    seMarker = peakMarkersHBC,
    ArchRProj = projPeaks,
    peakAnnotation = "Motif",
    cutOff = "FDR <= 0.05 & Log2FC >= 0.5"
  )
## ArchR logging to : ArchRLogs/ArchR-peakAnnoEnrichment-1f66690639ec-Date-2021-01-28_Time-23-37-49.log
## If there is an issue, please report to github with logFile!
## 2021-01-28 23:37:52 : Computing Enrichments 1 of 3, 0.059 mins elapsed.
## 2021-01-28 23:37:54 : Computing Enrichments 2 of 3, 0.083 mins elapsed.
## 2021-01-28 23:37:55 : Computing Enrichments 3 of 3, 0.102 mins elapsed.
## ArchR logging successful to : ArchRLogs/ArchR-peakAnnoEnrichment-1f66690639ec-Date-2021-01-28_Time-23-37-49.log
heatmapMotifsUp <- plotEnrichHeatmap(motifsUp, transpose = TRUE)
## ArchR logging to : ArchRLogs/ArchR-plotEnrichHeatmap-1f6617c4ead2-Date-2021-01-28_Time-23-37-56.log
## If there is an issue, please report to github with logFile!
## Adding Annotations..
## Preparing Main Heatmap..
pMotif <- ComplexHeatmap::draw(heatmapMotifsUp, heatmap_legend_side = "bot", annotation_legend_side = "bot")
pMotif

heatmapMotifsMatrix <- plotEnrichHeatmap(motifsUp, transpose = TRUE, returnMatrix = TRUE)
## ArchR logging to : ArchRLogs/ArchR-plotEnrichHeatmap-1f661cb734d9-Date-2021-01-28_Time-23-37-58.log
## If there is an issue, please report to github with logFile!
colnames(heatmapMotifsMatrix) <- unlist(lapply(strsplit(colnames(heatmapMotifsMatrix), split="_"), "[[", 1))

pMotif <- Heatmap(heatmapMotifsMatrix, 
        cluster_rows = FALSE,
        cluster_columns = FALSE,
        col = paletteContinuous(set = "comet", n = 100),
        column_names_gp = gpar(fontsize = 11),
        row_names_gp = gpar(fontsize = 13),
        heatmap_legend_param = list(title = "-log10 p-adj."))
pMotif


png("../plots/motifHeatmapHBCStates.png", width=10, height=7, units="in", res=300)
ComplexHeatmap::draw(heatmapMotifsUp, heatmap_legend_side = "bot", annotation_legend_side = "bot")
dev.off()
## quartz_off_screen 
##                 2

Export peak data for custom downstream analysis

useMatrix="PeakMatrix"
ArrowFiles <- getArrowFiles(projPeaks)
featureDF <- ArchR:::.getFeatureDF(head(ArrowFiles, 2), "PeakMatrix")
peakMat <- ArchR:::.getPartialMatrix(ArrowFiles, 
            featureDF = featureDF, threads = 1, useMatrix = "PeakMatrix", 
            cellNames = rownames(projPeaks@cellColData), progress = FALSE)
rownames(peakMat) <- featureDF$name
saveRDS(peakMat, file="../data/scATAC/peakMatArchrHbc_v2.rds")

Save ArchR project

saveArchRProject(ArchRProj = projPeaks, outputDirectory = "../data/scATAC/archR_samples_pairwise_v2", load = FALSE)

Composite plot

V1

library(cowplot)
## 
## ********************************************************
## Note: As of version 1.0.0, cowplot does not change the
##   default ggplot2 theme anymore. To recover the previous
##   behavior, execute:
##   theme_set(theme_cowplot())
## ********************************************************
library(grid)

# DR of all cells
pCellType

# DR of HBCs
pstate 

# Barplot of injured vs uninjured in HBC states
pBar

# Motif enrichment
ComplexHeatmap::draw(Heatmap(heatmapMotifsMatrix))

# Composite
cowplot::plot_grid(pCellType + scale_color_manual(values = RColorBrewer::brewer.pal(9, "Set1")[c(2, 5, 9)]),
                   pstate,
                   pBar + theme_bw() + ylab("Cell count") + xlab("Treatment"),
                   grid::grid.grabExpr(ComplexHeatmap::draw(pMotif)),
                   nrow=2,
                   ncol=2,
                   labels=letters[1:4])
ggsave("../plots/figure3.png", width=12, height=9)

V2

library(cowplot)
library(grid)

# DR of HBCs: treatment
pinj <- ggplot(umapDimsHBC, aes(x=UMAP1, y=UMAP2, color=treat)) +
  geom_point(size = 1, alpha=0.4) + 
  theme_classic() +
  labs(color = "Treatment") + 
  scale_color_manual(values = c("orange", "darkseagreen3")) 
  
# DR of HBCs: cell state
clHBCMerged2 <- as.character(clHBCMerged)
clHBCMerged2[clHBCMerged2 == "C1_Inj"] <- "Activated"
clHBCMerged2[clHBCMerged2 == "C2_Hybrid"] <- "Hybrid"
clHBCMerged2[clHBCMerged2 == "C3_UI"] <- "Resting"
pstate <- ggplot(umapDimsHBC, aes(x=UMAP1, y=UMAP2, color=clHBCMerged2)) +
  geom_point(size = 1, alpha=0.4) + 
  theme_classic() +
  labs(color = "Cell state")
 

# Barplot of injured vs uninjured in HBC states
df <- data.frame(cl=clHBCMerged2,
                 treatment=proj$treat)

pBar <- ggplot(df, aes(x=treatment, fill=treatment)) + 
  geom_bar(stat="count") +
  scale_fill_manual(values = c("orange", "darkseagreen3")) +
  facet_wrap(.~cl) +
  theme_classic()
pBar

# Motif enrichment
ComplexHeatmap::draw(Heatmap(heatmapMotifsMatrix))

# Composite
cowplot::plot_grid(pinj,
                   pstate,
                   pBar + theme_bw() + ylab("Cell count") + xlab("Treatment"),
                   grid::grid.grabExpr(ComplexHeatmap::draw(pMotif)),
                   nrow=2,
                   ncol=2,
                   labels=letters[1:4])
ggsave("../plots/figure3_v2.png", width=12, height=9)